Bacterial meningitis and pneumonia are responsible for a large worldwide disease burden, greater than other better-funded pathogenic diseases like HIV/AIDS and malaria. One of these pathogens, Haemophilus influenzae, inhabits the upper respiratory tract of most people, usually with no negative effect, but can cause several diseases-including meningitis, pneumonia, and ear infections-in infants, the elderly, and immune- compromised individuals. Like many other bacterial pathogens, H. influenzae can consume DNA by transporting it into their cells, breaking it down and reusing the subunits. But some of this DNA can recombine with a cell's own chromosome, interchanging versions of genes and adding or deleting genes. This natural competence mechanism allows related bacteria living in the same host to share genes, hastening their adaptation to host defenses and therapeutic interventions. The proposed research will link molecular studies of transformation with epidemiological studies of natural genetic diversity by experimentally measuring the chromosome-wide consequences of natural transformation. I will use the genomic DNA of clinical isolates to transform the chromosomes of competent recipient cells, and I will then apply massively parallel DNA sequencing to measure the extent and limits of natural transformation. I will use two complementary approaches: (1) measure the extent of transformation in a set of individual recombinant clones, and (2) measure the frequency of transformation across the chromosome from cultures of transformed cells. This research will be the first comprehensive experimental evaluation of genetic exchange in a human pathogenic bacterium (or any bacteria). PUBLIC HEALTH RELEVANCE: Understanding the limits of natural transformation has important public health value. Genetically heterogeneous bacteria in the same environment can share genes using the natural competence mechanism, offering a route for pathogens to exchange antibiotic resistance genes, virulence factors, and variants of antigenic surface markers. The explosion of genome sequences of naturally competent bacteria reveals extensive evidence of pervasive recombination between different pathogenic bacterial isolates, but the role of intrinsic biases in the natural transformation pathway for different DNA sequences has not been systematically addressed.